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Blood compatibility of biomaterials: further evaluation of the Lindholm test
Mason, R. G., B. A. Shinoda, et al. (1974), Biomater Med Devices Artif Organs 2(1): 21-30.

Blood compatibility of cetyl alcohol/polysorbate-based nanoparticles
Koziara, J. M., J. J. Oh, et al. (2005), Pharm Res 22(11): 1821-8.
Abstract: PURPOSE: Pegylated and nonpegylated cetyl alcohol/polysorbate nanoparticles (E78 NPs) are being tested as drug carriers for specific tumor and brain targeting. Because these nanoparticle formulations are designed for systemic administration, it is important to test the compatibility of these lipid-based NPs with blood and blood cells. METHODS: The hemocompatibility of E78 NPs was evaluated with a particular focus on hemolytic activity, platelet function, and blood coagulation. Human red blood cell lysis was determined by measuring hemoglobin release. Activation and aggregation of human platelets were determined using flow cytometry and aggregometry, respectively. Finally, the whole blood clotting time was measured using human blood. RESULTS: E78 NPs did not cause in vitro red blood cell lysis at concentrations up to 1 mg/mL. In addition, under conditions tested, E78 and polyethylene glycol (PEG)-coated E78 NPs (PEG-E78 NPs) did not activate platelets. In fact, both NP formulations very rapidly inhibited agonist-induced platelet activation and aggregation in a dose-dependent manner. Additionally, E78 NPs significantly prolonged in vitro whole blood clotting time at a concentration of 500 microg/mL or greater. CONCLUSIONS: It was concluded that PEG-coated and nonpegylated E78 NPs have potential blood compatibility at clinically relevant doses. Based on the calculated nanoparticle-to-platelet ratio, the concentration at which E78 NPs could potentially affect platelet function in vivo was approximately 1 mg/mL.

Blood compatibility of thermoplastic polyurethane membrane immobilized with water-soluble chitosan/dextran sulfate
Lin, W. C., D. G. Yu, et al. (2005), Colloids Surf B Biointerfaces 44(2-3): 82-92.
Abstract: Water-soluble chitosan (WSC)/dextran sulfate (DS) was immobilized onto the surface of thermoplastic polyurethane (TPU) membrane after ozone-induced graft polymerization of poly(acrylic acid) (PAA). The surface was characterized with contact angle measurement and X-ray photoelectron spectroscopy (XPS). The adsorption of human plasma fibrinogen (HPF) followed the Langmuir adsorption isotherm. The results showed that the surface density of peroxides generated and poly(acrylic acid) (PAA) grafted reached the maximum value at 20 min of ozone treatment. It was found that the WSC- and DS-immobilized amount increased with pH and the molecular weight of WSC. The membrane/water interfacial free energy increased with PAA-grafting and WSC/DS-immobilization, indicating the increasing wettability of TPU membrane. The adsorption of HPF on TPU-WSC/DS membranes could be effectively curtailed and exhibited unfavorable adsorption. Moreover, WSC/DS immobilization could effectively reduce platelet adhesion and prolong the blood coagulation time, thereby membrane improving blood compatibility of TPU membrane. In addition, the in vitro cytotoxicity test of PEC modification was non-cytotoxic according to much low growth inhibition of L929 fibroblasts. Furthermore, TPU-WSC/DS membranes exhibited higher cell viability than native TPU membrane.

Blood compatible polymers in intensive care units: state of the art and current aspects of biomaterials research
Ragaller, M., C. Werner, et al. (1998), Kidney Int Suppl 64: S84-90.
Abstract: The use of artificial organs in cases of acute renal failure, acute respiratory distress syndrome (ARDS) and multiorgan dysfunction syndrome (MODS) has lead to a significant reduction of mortality. However, the interaction between body and biomaterials results in the activation of the coagulation system and in the induction of systemic inflammatory response syndrome. The necessary anticoagulation may be contraindicated and may even further increase the risk for the patient. This article evaluates the currently applied polymeric materials used in intensive care units (ICU) and gives a possible outlook into future developments. It is emphasized that systematic interdisciplinary research of physicians and biomaterial scientists is essential for the successful development of new polymers with improved biocompatibility. For this purpose a brief overview of analytical techniques for surface characterization is given, and future developments to a fully biocompatible polymer are described.

Blood proteins do not promote adherence of coagulase-negative staphylococci to biomaterials
Muller, E., S. Takeda, et al. (1991), Infect Immun 59(9): 3323-6.
Abstract: We studied the effects of in vitro and in vivo coating of catheters with human blood proteins on binding of coagulase-negative staphylococci. Coating resulted in no enhancement of binding. Catheters coated in vitro bound fewer organisms than uncoated catheters. Host proteins do not enhance adherence of coagulase-negative staphylococci to biomaterials.

Blood-biomaterial interactions in a flow system in the presence of bacteria: effect of protein adsorption
Sapatnekar, S., K. M. Kieswetter, et al. (1995), J Biomed Mater Res 29(2): 247-56.
Abstract: An in vitro continuous flow system with whole human blood was used to study blood-biomaterial interactions on a base polyurethane and three modified surfaces in the presence and absence of circulating Staphylococcus epidermidis. We hypothesized that the composition of the protein layer adsorbed on the surface of the biomaterial would influence the response of blood components and bacteria. We examined the test surfaces for adsorption of nine plasma proteins and adsorption profiles differed on the four surfaces. The positively charged surface, UC, adsorbed significantly higher amounts of fibronectin (P <.01), von Willebrand factor (P <.01), and fibrinogen (P <.05) than the other materials. As a consequence of increased adsorption of these adhesive proteins, the adhesion of platelets and bacteria was greater on UC than on any other surface. On the base polyurethane, BC, and the negatively charged surface, UA, protein adsorption was low, and these materials were largely free of adherent blood cells and bacteria. The heparinized surface, UH, adsorbed higher quantities (P <.01) of Hageman factor and high molecular weight kininogen relative to the other surfaces. Platelet adhesion, and surface coagulation were prominent on UC, and may have contributed to increased bacterial adhesion on this surface. In the presence of circulating bacteria, adsorption was generally lower than in the absence of bacteria. The pattern of protein adsorption was largely unaffected by the strain of circulating bacteria, but platelet responses (adhesion and activation) were greater in the presence of slime-producing S. epidermidis as compared to the non-slime-producing strain, suggesting that slime may have a direct activating effect on platelets.

Blood-compatible biomaterials by surface coating with a novel antithrombin-heparin covalent complex
Klement, P., Y. J. Du, et al. (2002), Biomaterials 23(2): 527-35.
Abstract: Covalent antithrombin-heparin complex (ATH) was covalently grafted to a polycarbonate urethane (Corethane) endoluminal graft (a kind gift of Corvita Corporation) after being activated using 0.3% m/m NaOCl in 0.15 M phosphate pH 6.0. ATH graft density (1.98 x 10(-7) mol/m2) was 6 times the maximum amount of unfractionated heparin (UFH) that could be bound to polycarbonate urethane surfaces. Surface-bound ATH could be stored in sterile 0.15 M NaCl at 4 degrees C for at least 2 months with good antithrombotic activity before being implanted into rabbits. Analysis of ATH-coated tubing showed that it contained significant direct thrombin inhibitory activity. In vivo testing in a rabbit model was compared to non-activated non-coated surfaces, activated-non-coated surfaces, hirudin-coated surfaces and antithrombin (AT)-coated surfaces. The weight of the clot generated in the ATH-coated graft tubing was significantly less than the weight of the clot generated within the hirudin-coated graft (p = 0.03 with a 1-tailed Student's t test). The anticoagulant nature of ATH grafts in vivo was shown to be due to bound ATH because boththe AT-coated surfaces and non-coated but activated surfaces showed similar thromboresistant efficacy to that of untreated material (ANOVA; p < 0.05). Apart from the direct antithrombin activity that contributed to much of the prolonged patency in vivo, surface-bound ATH likely catalyzed AT inhibition of thrombin, as evidenced by a significant number of 125I-AT binding sites (> or = 1.5 x 10(-8) mol/m2). Thus, ATH appears to be a good candidate for coating cardiovascular devices, such as endoluminal grafts, with high levels of substitution and significant long-term blood-compatibility.

Blood-contacting biomaterials: bioengineering viewpoints
Courtney, J. M., N. M. Lamba, et al. (1995), Artif Organs 19(8): 852-6.
Abstract: The investigation of blood-contacting biomaterials is an important challenge and is relevant for an improvement in the clinical application of biomaterials. With the purpose of improved clinical treatment, bioengineering viewpoints of blood-contacting biomaterials cover the material options and selection, the utilization of materials, the development of materials with better properties, and processing characteristics, and the design of relevant evaluation procedures. The bioengineering objective remains that of achieving an enhanced understanding of the relationship between a biomaterial and the biological response.

Blood-filled spaces with and without deproteinized bone grafts in guided bone regeneration. A histomorphometric study of the rabbit skull using non-resorbable membrane
Okazaki, K., Y. Shimizu, et al. (2005), Clin Oral Implants Res 16(2): 236-43.
Abstract: This experimental study evaluated the effects of deproteinized bone grafts on guided bone regeneration (GBR). A groove was made in the bone marrow of the external cortical plate of the skull. A dome of non-resorbable membrane was placed on the groove and secured with titanium pins. The secluded graft space was filled with autogenous blood clots (control group) and deproteinized bone particles (experimental group). The rabbits were sacrificed 2, 4, 8 and 12 weeks after the operation. Decalcified and paraffin-embedded, transverse 3-mum-thick sections were made and stained with hematoxylin and eosin. The proportions of newly formed bone and newly formed bone-graft particle contact surfaces were histomorphometrically measured in the basal, central, and peripheral areas from the cortical plate to the top of the dome. In the control group, the basal area showed a significant increase at 4 weeks (P<0.01) and a significant decrease at 8 weeks (P<0.01). The central and peripheral areas showed gradual increases in the proportion of newly formed bone. The experimental group showed significant increase at 4 weeks in the basal area and at 8 weeks in central and peripheral area (P<0.01). There were significant differences between both groups in basal and central area (P<0.01). The proportion of newly formed bone-graft particle contact length showed significant increases at 4 weeks (P<0.01) and no significant decreases at 8 and 12 weeks in three areas. The present study showed that deproteinized bone grafts maintain newly formed bone in extensive areas for a prolonged period during GBR.

BMP-2 liberated from biomimetic implant coatings induces and sustains direct ossification in an ectopic rat model
Liu, Y., K. de Groot, et al. (2005), Bone 36(5): 745-57.
Abstract: INTRODUCTION: Using a rat model, we evaluated the kinetics and histomorphometry of ectopic bone formation in association with biomimetic implant coatings containing BMP-2. MATERIALS AND METHODS: One experimental and three control groups were set up: titanium-alloy discs coated with a biomimetically co-precipitated layer of calcium phosphate and BMP-2 [1.7 microg per disc (incorporated-BMP group)]; uncoated discs (control); discs biomimetically coated with a layer of calcium phosphate alone (control); and discs biomimetically coated with a layer of calcium phosphate bearing superficially adsorbed BMP-2 [0.98 microg per disc (control)]. Discs (n = 6 per group) were implanted subcutaneously in rats and retrieved at 7-day intervals over a period of 5 weeks for kinetic, histomorphometrical, morphological and histochemical analyses. RESULTS: In the incorporated-BMP-2 group, osteogenic activity was first observed 2 weeks after implantation and thereafter continued unabated until the end of the monitoring period. The net weekly rates of bone formation per disc were 5.8 mm3 at 2 weeks and 3.64 mm3 at 5 weeks. The total volumes of bone formed per disc at these junctures were 5.8 mm3 and 10.3 mm3, respectively. Bone tissue, which was formed by a direct ossification mechanism, was deposited at distances of up to 340 microm from the implant surfaces. The biomimetic coatings were degraded gradually, initially by foreign body giant cells alone and then also by osteoclasts. Forty percent of the coating material (and thus presumably of the incorporated BMP-2) remained at the end of the monitoring period. Hence, 60% of the incorporated BMP-2 had been released. At this 5-week juncture, no bone tissue was associated with any of the control implants. CONCLUSION: BMP-2 incorporated into biomimetic calcium phosphate coatings is capable not only of inducing bone formation at an ectopic site in vivo but also of doing so with a very high potency at a low pharmacological level, and of sustaining this activity for a considerable period of time. The sustainment of osteogenic activity is of great clinical importance for the osseointegration of dental and orthopedic implants.

Bond strength and SEM observation of CO2 laser irradiated dentin, bonded with simplified-step adhesives
Koshiro, K., S. Inoue, et al. (2005), Oper Dent 30(2): 170-9.
Abstract: This study investigated, mechanically and morphologically, whether the dentin surface irradiated by CO2 laser could be a possible adherent when bonded with simplified-step adhesives. Buccal enamel and cementum of extracted human premolars were removed to expose a flat dentin surface. The dentin surfaces were irradiated continuously with CO2 laser at 1.0 W. Before bonding with either a single-bottle adhesive (Single Bond) or a self-etching priming system (Mega Bond), the irradiated dentin surface was treated as follows: no treatment, NaHCO3 powder abrasion and wet-grinding with 600-grit SiC paper. The treated dentin surfaces were bonded to resin composite with either of the two adhesives. Non-irradiated dentin surfaces were also used as control. Resin bonded specimens were stored in water at 37 degrees C for 24 hours and subjected to microtensile bond test. Additionally, to observe the resin/irradiated dentin interface, resin-bonded specimens were similarly prepared, sectioned into slabs, embedded in epoxy resin, polished with diamond pastes, sputter coated Au-Pd and examined with scanning electron microscopy (SEM). After SEM observation, the specimens were further polished with diamond paste to remove the Au-Pd sputter-coat, immersed in HCL and NaOCl and finally observed by SEM again. In the presence of carbonized dentin, microtensile bond strength drastically decreased but recovered to the control value by removing the carbonized dentin layer visually with SiC paper for both adhesive systems. However, the laser-affected dentin that remained on the bonded interface was easily dissolved with NaOCl and HCl.

Bone challenges for the hand surgeon: from basic bone biology to future clinical applications
Weyand, B. and H. P. von Schroeder (2005), Clin Plast Surg 32(4): 537-47, vii.
Abstract: Bone is a complex tissue composed of a calcified extracellular matrix with specialized cells that produce, maintain, and resorb the bone. Bone also has a rich vascular and neural supply. Bone has a great capability of regeneration, healing, and remodelling that is influenced by external factors, such as stress forces, and internal regulators that include hormones, vitamins, and growth factors. These factors dictate bone biology, and variations result in pathophysiologic conditions that have clinical implications in hand surgery. Solutions to the challenges in hand surgery rely on a thorough understanding of the biology of bone.

Bone defects of the facial skeleton - replacement with biomaterials
Zboril, V., J. Pazdera, et al. (2003), Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub 147(1): 51-6.
Abstract: The authors report on their experience with bone defect treatment following surgery of jaw-bone cysts. This is based on the use of cadaveric ground spongiosis saturated with tetracycline and metronidazol solution. The study shows the above mentioned material is very suitable for bone defect therapy, and cost effective. The results were verified by clinical and X-ray examination.

Bone formation and resorption of highly purified beta-tricalcium phosphate in the rat femoral condyle
Kondo, N., A. Ogose, et al. (2005), Biomaterials 26(28): 5600-8.
Abstract: The aim of this study was to examine the chronological histology associated with highly purified beta-tricalcium phosphate (beta-TCP) implanted in the rat femoral condyle. Specimens were harvested on days 4, 7, 14, 28 and 56 after implantation, and were analyzed by tartrate-resistant acid phosphatase (TRAP) staining, immunohistochemistry of the ED1 protein as a marker of the phagocyte system, and in situ hybridization with digoxigenin-labeled alpha1 chain of type I procollagen (COL1A1), osteopontin and osteocalcin. beta-TCP was resorbed in a chronological manner. Although new bone was not observed on day 4, fibroblast-like cells around beta-TCP were positive for COL1A1 and osteopontin mRNA. New bone formation presented after day 7. In the double-staining for OPN and ED1 on day 7, most cells around beta-TCP were positive for either osteopontin mRNA or ED1 protein. However, there were some doubly positive multinucleated cells, suggesting that they belonged to the mononuclear phagocyte system. After day 28, the implanted region was replaced with bone marrow. Multinucleated TRAP-positive and ED1-positive cells which adhered to beta-TCP at all stages seemed to be osteoclasts and they continuously resorbed beta-TCP. beta-TCP has a good biocompatibility since both bioresorption and bone formation started at an early stage after implantation.

Bone formation at titanium implants prepared with iso- and anisotropic surfaces of similar roughness: an in vivo study
Goransson, A. and A. Wennerberg (2005), Clin Implant Dent Relat Res 7(1): 17-23.
Abstract: BACKGROUND: Implant surface topography influences the bone response after implantation. However, the importance of surface orientation is not known. Purpose: The aim of this study was to investigate the bone tissue response and the stability of titanium implants prepared with isotropic and anisotropic surfaces of similar roughness. MATERIALS AND METHODS: A total of 18 implants were divided into two groups and were inserted into the femurs of nine rabbits for 12 weeks. Confocal laser scanning microscopy was used for the topographic description to verify that the two different surfaces were modified as intended. The stability of the implants was recorded by resonance frequency (RF) measurements at insertion and at time of removal, after which the implants were evaluated histomorphometrically. RESULTS: RF measurements showed that implant stability increased with time. However, there was no significant difference between the two different surface modifications at insertion and after 12 weeks. The histomorphometric comparison revealed no statistically significant differences in regard to either bone-to-metal contact or bone area inside the threads. CONCLUSION: Titanium implants prepared with isotropic and anisotropic surfaces of similar roughness integrate similarly to bone during the 3 months after implantation.

Bone formation by enamel matrix proteins and xenografts: an experimental study in the rat ramus
Donos, N., D. Bosshardt, et al. (2005), Clin Oral Implants Res 16(2): 140-6.
Abstract: The aim of this study was to evaluate whether the use of enamel matrix proteins with or without the use of deproteinized bovine bone influences bone formation when used as an adjunct to guided bone regeneration (GBR). Twenty rats, divided into four groups of five animals each, were used in this study. Group A1: A hemispherical PTFE capsule was placed empty on the lateral aspect of the mandibular ramus (GBR). At the contralateral side of the jaw, the capsule was filled with an enamel matrix derivative (EMD) before its placement. The healing period was 60 days. Group A2: The animals were treated in the same manner as in Group A1 but with a healing period of 120 days. Group B1: The animals were treated in the same manner as in Group A1 with the difference that deproteinized bovine bone mineral (DBBM) particles were packed in the capsule. At the contralateral side of the jaw, the capsule was filled with a mixture of EMD and DBBM. The healing period was 60 days. Group B2: The same treatment as in B1 but with a healing period of 120 days. The histological analysis revealed that in Groups A1 and A2 newly formed bone was covering a significant part of the empty capsules (GBR). The use of EMD in the capsule did not offer any added benefit to the use of the capsule alone in terms of new bone formation. At Groups B1 and B2, the presence of DBBM and/or EMD did not positively affect the amount of new bone formation. It can be suggested that neither the application of EMD nor the use of DBBM or the combination of EMD and DBBM results in enhanced amounts of bone formation in comparison with the GBR procedure alone.

Bone formation in a long bone defect model using a platelet-rich plasma-loaded collagen scaffold
Sarkar, M. R., P. Augat, et al. (2006), Biomaterials 27(9): 1817-23.
Abstract: Platelet-rich plasma (PRP), a platelet concentrate made of autogenous blood, has been used in recent years to improve bone defect healing particularly in maxillofacial reconstructions. The aim of the present study was to assess the effect of PRP on new bone formation in a critical diaphyseal long bone defect. A critical size defect (2.5cm) in the tibial diaphysis of 16 sheep was supplied either with autogenous PRP in a collagen carrier or with collagen alone (controls). Platelets were enriched about 3.5 fold compared to normal blood in the PRP. After 12 weeks, the explanted bone specimens were quantitatively assessed by X-ray, computed tomography (CT), biomechanical testing and histological evaluation. Bone volume, mineral density, mechanical rigidity and histology of the newly formed bone in the defect did not differ significantly between the PRP treated and the control group, and no effect of PRP upon bone formation was observed. It was suggested that PRP does not enhance new bone formation in a critical size defect with a low regenerative potential. Such bone defects might require more potent stimuli, e.g. combinations of functional biomaterials or autografts, precursor cells or osteoinductive growth factors.

Bone formation in beta-tricalcium phosphate-filled bone defects of the rat femur: morphometric analysis and expression of bone related protein mRNA
Shiratori, K., K. Matsuzaka, et al. (2005), Biomed Res 26(2): 51-9.
Abstract: The purpose of the current study was to evaluate the bone formation when beta-tricalcium phosphate (TCP) was implanted in bone defects of rat femurs. beta-TCP granules were applied to defects created in the femurs of 65 male rats who were sacrificed 3, 7, 10, 14 or 30 days later. Bone tissues were embedded in paraffin, serial sections were cut and then stained with hematoxylin-eosin. Histomorphometric analyses were also conducted. Furthermore, total mRNAs were extracted, homogenized, and reverse transcribed, after which quantitative PCR assays were conducted with a LightCycler using the double-stranded DNA dye Syber Green I with primers for either rat osteopontin or osteocalcin. Tissues in defects without beta-TCP were used as controls. The amount of newly formed bone tissue in the beta-TCP implanted group was significantly greater in both the side areas and the central area of defects than in the control group. Expressions of osteopontin and osteocalcin mRNAs of cells in the defects of the experimental group were up-regulated compared with the control group at all time periods. Taken together, these results prove that beta-TCP is an appropriate material for osteoconduction and promotes bone formation in bone defects.

Bone formation in calvarial defects of Sprague-Dawley rats by transplantation of calcium phosphate glass
Moon, H. J., K. N. Kim, et al. (2005), J Biomed Mater Res A 74(3): 497-502.
Abstract: The purpose of this study was to investigate the bone-regenerative effect of calcium phosphate glass in vivo. We prepared amorphous calcium phosphate glass powder having a mean particle size of 400 microm in the system CaO-CaF2-P2O5-MgO-ZnO. Calvarial critical-sized defects (8 mm) were created in 60 male Sprague-Dawley rats. The animals were divided into an experimental group and control group of 30 animals each. Each defect was filled with a constant weight of 0.5 g calcium phosphate glass powder mixed with saline. As a control, the defect was left empty. The rats were sacrificed 2, 4, or 8 weeks postsurgery, and the results evaluated using radiodensitometric and histological studies; they were also examined histomorphometrically. When the calcium phosphate glass powders with 400-microm particles were grafted, the defects were nearly completely filled with new-formed bone in a clean healing condition after 8 weeks. It was observed that the prepared calcium phosphate glass enhanced new bone formation in the calvarial defect of Sprague-Dawley rats and could be expected to have potential for use as a hard tissue regeneration material.

Bone graft alternatives in the treatment of benign bone tumors
Rougraff, B. T. (2005), Instr Course Lect 54: 505-12.
Abstract: Most bone grafting procedures are done during spinal fusion and to treat patients with skeletal trauma. Very few studies have addressed the bone grafting of skeletal defects after benign bone tumor excision. Contained defects have been treated with autogenous bone grafts, fresh-frozen allografts, freeze-dried allografts, demineralized bone matrix, and ceramic materials. Additionally, bone morphogenetic proteins may provide a future treatment option for bone tumor reconstruction.

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